AC is a 12-transmembrane protein that catalyzes the conversion of ATP to cAMP upon the stimulation of various G-protein coupled receptors such as β-adrenergic receptor (β-AR). Nine mammalian AC subtypes have been identified, and each subtype shows distinct tissue distributions, and biological and pharmacological properties (Iwatsubo et al., Endocr Metab Immune Disord Drug Targets. September 2006;6(3):239-247). Stimulation of G protein-coupled receptors induces binding of the stimulatory Gα subunit (Gsα) to AC, and enhances its catalytic activity to convert ATP into cAMP. cAMP regulates multiple downstream molecules, via protein kinase A (PKA) and exchange protein activated by cAMP (Epac).
AC5 is a major cardiac subtype of AC, which provides 20% of total AC activity in the heart, and recent studies including ours revealed its crucial role in progression of HF (Iwatsubo et al., J Biol Chem. Sep. 24, 2004;279(39):40938-40945; Okumura et al., Circ Res. Aug. 22, 2003 2003;93(4):364-371). AC5KO mice showed decreased myocardial apoptosis and preserved cardiac function in HF models induced by chronic pressure overload (Okumura et al. Proceedings of the National Academy of Sciences. 2003;100(17):9986-9990), chronic β-AR stimulation (Okumura et al., Circulation. 2007;116(16):1776-1783) and aging (Yan et al., Cell. Jul. 27, 2007;130(2):247-258). In all these HF models, myocardial apoptosis, which is a major cause for progression of HF, was significantly decreased in AC5KO, indicating that AC5 plays a central role in inducing apoptosis and subsequent development of HF. Moreover, AC5Tg showed decreased left ventricular ejection fraction (LVEF) and increased apoptosis in response to chronic pressure overload, indicating that AC5 accelerates the progression of HF by inducing myocardial apoptosis. These data strongly suggest that among mechanisms by which myocardial apoptosis occurs such as renin-angiotensin-aldosterone, death receptor and calcium signaling, sympathetic activity overdrive, particularly via stimulating AC5, plays a major role in inducing myocardial apoptosis and development of HF.
Classic inhibitors of AC, known as P-site inhibitors, have been studied since the 1970's. It was first thought that there was an adenosine-reactive site within intracellular domain of AC, the “P” site, which inhibits the catalytic activity of AC. In spite of their similar chemical structure to the substrate ATP, P-site inhibitors showed un- or non-competitive inhibition with respect to ATP, indicating little influence on molecules which have ATP-binding site (Londos et al., Proc Natl Acad Sci U S A. December 1977;74(12):5482-5486). Although it has been a very attractive idea to develop P-site inhibitors with enhanced AC subtype selectivity, few attempts have been successful due to the difficulties of experiments in which the selectivity of each AC isoforms can be examined in vitro. However, several groups including ours have developed such experimental systems using the baculovirus-based recombinant AC overexpression system (Iwatsubo et al., J Biol Chem. Sep. 24, 2004;279(39):40938-40945; Onda et al. J Biol Chem. Dec. 21, 2001;276(51):47785-47793).
9-β-D-arabinofuranosyladenine (AraAde) contains an adenosine-like structure where the adenine ring is essential not only for binding to the AC catalytic core but also for penetrating the plasma membrane (Iwatsubo et al. J Biol Chem. 2004;279(39):40938-40945, Onda et al. J Biol Chem. 2001;276(51):47785-47793. Tesmer et al. Biochemistry.2000;39(47):14464-14471. Tesmer et al. Science. 1999;285(5428):756-760). For example, NKY80, which does not contain adenosine within its structure, showed moderate inhibition of purified AC5 protein in vitro, but it does not inhibit cAMP accumulation in cultured cardiac myocytes, indicating that the adenosine structure seems essential for penetrating the plasma membrane (Iwatsubo, et al. J Biol Chem.279(39):40938-40945). In addition, adenosine hardly crosses through the blood-brain barrier (BBB) (Isakovic et al. Journal of Neurochemistry, 90(2):272-286.), having little influence on brain function; this is important because AC5 is also expressed in the striatum other than the heart, thus by passing BBB AC5 inhibitors may cause adverse effects in the brain.
Obesity is a disease in which excess body fat has accumulated to such an extent that health may be negatively affected. Many studies have shown an association between excessive body weight and various diseases, particularly cardiovascular diseases, diabetes mellitus type 2, sleep apnea, certain types of cancer, and osteoarthritis. As a result, obesity has been found to reduce life expectancy. Obesity is a global epidemic. As of 2005 the World Health Organization estimates that at least 400 million adults (9.8%) are obese. The United States has the highest rates of obesity in the developed world. It was reported in 2005 that about 119 million, or 64.5%, of US adults are either overweight or obese. Obesity is a public health problem because of its prevalence, costs and burdens. Obesity can be caused by dietary, behavioral and genetic factors. There have been several genes identified that are involved in the development of obesity.
Obesity is a complex disease influenced by diet, exercise, and a complex biology. Many peer-reviewed studies show that people who successfully complete weight loss program generally regain weight. Bariatric surgery to reduce the size of the stomach (gastric bypass surgery) is the only effective treatment for causing weight loss in morbid obese people. Two prescription drugs have been approved by the Food and Drug Administration (FDA) for long-term weight loss (Sibutramine and Orlistat, combined market $500 million). These drugs work in different ways, cause different side effects and only result in modest weight loss. Thus, discovery of novel obesity treatments is urgently needed to treat this epidemic.
Obesity in humans and in rodents is usually associated with high circulating leptin levels and leptin resistance. Leptin, the protein product of the ob gene, is predominantly secreted from white adipose tissue, and acts on the brain to regulate food in-take, energy expenditure, and neuroendocrine function. In obese (ob/ob) mice that lack functional leptin, recombinant leptin is highly effective at reversing obesity. However, most cases of obesity in rodents and humans are associated with high circulating leptin levels; the resistance to leptin that characterizes these states has not yet been explained. Potential sites for leptin resistance include the blood-brain-barrier transport system and the leptin signaling mechanism in leptin-responsive neurons in the hypothalamus. Rodents with obesity induced by high-fat diet (diet-induced obesity, or DIO) become hyperleptinemic, and food intake and body weight are resistant to the effects of exogenous leptin administered peripherally.
Diabetes mellitus induces a variety of metabolic abnormalities because of insufficient insulin action. Of these, abnormalities in glucose metabolism are the most specific and are manifested clinically as hyperglycemia after glucose ingestion. In type 2 diabetes mellitus, which affects the majority of patients with diabetes mellitus, the factors involved in the pathogenesis and the progression of the disease are insufficient insulin secretion and decreased insulin sensitivity (insulin resistance). The relationship between insufficient insulin secretion and diabetes mellitus was underscored by the discovery of the causative gene for maturity onset diabetes of the young, and abnormalities in insulin secretion are considered to be particularly important. Prevention of the progression of pancreatic β-cell dysfunction in subjects with diabetes mellitus should be a key in the long-term management of this disease.
Diabetes mellitus is a major risk factor of HF and coronary artery disease (Fuller, et al., Br Med J (Clin Res Ed) 1983, 287(6396): 867-870). Diabetes mellitus induces a variety of metabolic abnormalities because of insufficient insulin action. Of these, abnormalities in glucose metabolism are the most specific and are manifested clinically as hyperglycemia after glucose ingestion. In type 2 diabetes mellitus, the factors involved in the pathogenesis and the progression of the disease are insufficient insulin secretion and decreased insulin sensitivity (insulin resistance).
Exercise intolerance is a common feature in patients with HF and affects their quality of life, thus becoming an important target for therapies. In addition, exercise training has been widely understood as effective treatment for HF (Baladay, Ann Med, 1998, 30 Suppl 1:61-65; Massie, Am J Med., 1988, 84(3A): 75-82). However, paradoxically, this training is prevented by exercise intolerance arising from HF. Therefore, increasing exercise capacity is favorable for HF patients in terms of not only improving prognosis and quality of life but also providing effective therapy for HF.
Hyperglycemia and hyperlipidemia have been shown to affect the mechanism of insulin secretion. Impaired glucose-mediated insulin secretion from pancreatic β-cells leads to insulin insufficiency and thus hyperglycemia and lipid metabolism abnormalities. The hyperglycemic state leads to overworking of pancreatic β-cells and a decreased ability to secrete insulin. C57BL/KsJ db/db mice that have the db mutation exhibit a severe insulin resistance and impaired insulin secretion. They are widely used as experimental models of obese type 2 diabetes mellitus. Briefly, after birth these mice have unrepressed eating behavior, become obese, and develop severe insulin resistance associated with hyperinsulinemia, hyperglycemia, and hypertriglyceridemia, so that by 3-6 months after birth, the pancreatic islet β-cells reduce their mass, resulting in severe insufficiency of insulin secretion.
Obesity, diabetes and exercise are inextricably linked such that an agent that improves exercise tolerance or prevents obesity or the development of diabetes will likely have a therapeutic role in all three conditions. Obesity, a global epidemic, promotes diabetes and is a major cardiovascular risk factor, resulting in reduced life expectancy. Finding a novel therapeutic approach would be a major advance. Recently, a novel, genetically engineered mouse model, where the adenylyl cyclase (AC) type 5 isoform is knocked out (AC5 KO) was reported. AC5 inactivation resulted in increased longevity and was protective against stress. Furthermore, the AC5 KO mice ate more than WT mice, but weighed less, suggesting that AC5 inhibition could be a novel approach to weight loss. The AC5 KO mouse also demonstrates enhanced exercise tolerance. A pharmacological AC5 inhibitor protects against cardiovascular stress.